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OBJECTIVE This work aimed to investigate the anti-rheumatoid arthritic effect of gentio-picroside from Gentiana macrophylla Pall using an animal model of adjuvant induced arthritis. METH-ODS Adjuvant arthritis was induced in fifty SD male rats,which were randomly divided into five groups (n=10):control(0.5% CMC-Na)group,AIA(rats with CFA)group,dexamethasone(1 mg·kg-1)group, gentiopicroside(50 mg·kg-1)group,and gentiopicroside(100 mg·kg-1)group.Rats were administered intragastrically with drugs or CMC-Na once a day for a period of 2 weeks.Paw swelling,arthritic index, histological changes were assessed to evaluate the anti-arthritic effect.Weight growth,spleen and thymus indexes were also investigated in.RESULTS Gentiopicroside at dose of 100 mg·kg-1significantly inhibited the secondary paw swelling(P<0.05)and arthritis index(P<0.05),decreased synovial inflammatory infil-tration, synovial hyperplasia and bone erosion. Furthermore, gentiopicroside showed no immunosup-pressive adverse effects in body weight, index of spleen and thyums compared with dexamethasone administration (P<0.05, P<0.01). CONCLUSION Gentiopicroside possessed anti-arthritic efficacy in AIA rats without immunosuppressive effects.
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Objective: An ultra performance liquid chromatographic (UPLC) method was developed for simultaneously determining seven constituents, such as loganic acid, swertiamarin, 6'-O-β-D-glucosyl gentiopicroside, gentiopicroside, sweroside, isoorientin, and isovitexin, from the medicinal plants of Gentiana macrophylla. Methods: The separation was performed on an Acquity UPLC® BEH C18 column (50 mm ×2.1 mm, 1.7 μm) through a gradient elution of methanol-0.04% aqueous phosphorite at a flow rate of 0.3 mL/min. The detection wavelength was 242 nm, and the column temperature was set at 30℃. Results: For the seven analytes, loganic acid, swertiamarin, 6'-O-β-D-glucosyl gentiopicroside, gentiopicroside, sweroside, isoorientin, and sovitexin, a good linearity (r≥0.9995) was obtained in the range of 2.100-537.100, 1.050-270.000, 0.920-236.000, 11.100-2830.000, 0.750-192.000, 0.167-102.000, and 0.216-52.800 μg, respectively. Their average recoveries (n=6) were 97.83%-100.08%, respevtively, with RSD values less than or equal to 3.76%. Conclusion: The UPLC method is simpler and more effective than HPLC, and can be used for the simultaneous determination of seven indicative constituents in medicinal plants of G. macrophylla.
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Objective: To clone the gene sequence of the key enzyme in synthesis pathway of terpenoids, 1-deoxy-D-xylulose 5-phosphate reductoisomerase (DXR) from Gentiana macrophylla (GmDXR) and to analyze its characteristic and expression. Methods: GmDXR gene was cloned and the sequence characterization was analyzed with bioinformatics methods, then the expression patterns of GmDXR were studied by real-time PCR. Results: GmDXR gene contained a completed open reading frame of 1 428 bp encoding 475 amino acids, GmDXR has high homology (≥85%) with DXR proteins from Rauvolfia verticillata, Lycopersicon esculentum, and other plants. Further analysis showed that GmDXR had non-transmembrane domain structure and signal peptide, which mainly located in the chloroplast. Quantitative PCR results showed that GmDXR mainly expressed in the leaves of G. macrophylla and could be induced by plant hormone such as methyl jasmonate (MeJA). Conclusion: GmDXR contains conserved structures of DXR protein. GmDXR expresses variously in different organs of G. macrophylla and could be induced by MeJA. It is very helpful for the future research on biosynthetic mechanism of iridoid compounds in G. macrophylla.
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Objective To investigate the effects of carbohydrate and its concentration,nitrogen source,and metal ions on suspension cell growth of Gentiana macrophylla and gentiopicroside accumulation. Methods The cell dry weight and gentiopicroside content were measured under factors of different carbohydrate,NH4+/NO3-,Ca2+,and Zn2+ at various concentration. Results The optimum culture conditions are as follows:3% sucrose,NH4+/NO3-of 20/40,3 mmol/L Ca2+,0.007 49 mmol/L Zn2+. Conclusion Different carbohydrates and their concentration have significant effects on cell growth and gentiopicroside accumulation while high concentration of carbohydrates does not have such effect. NO3-is better than NH4+ for the cell growth. Ca2+ Remarkably influences both cell growth and gentiopicroside accumulation while Zn2+ contributes only to gentiopicroside content.
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Objective To establish cell suspension culture system of Gentiana macrophylla.Methods Good cell line was selected by the methods of small cell aggregate and the effects of different media,inoculation age of cells,initial pH value,culture temperature,and rotation speed on cell growth and gentiopicroside accumulation were investigated.Results The eligible cell suspension culture conditions were:MS media as the basic medium,cells with inoculation age of 15 d,initial pH 7,and rotation speed of 110 r/min at 25 ℃,culturing in Erlenmeyer flasks.Conclusion Optimal cell suspension culture system of G.macrophylla has been established by shake flasks.The different effects of media on cell growth and gentiopicroside accumulation indicate that some inorganic ions have significant effects on regulation.
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Objective To systematically study the chemical constituents in the roots of Gentiana macrophylla from Shaanxi Province. Methods Various column chromatographic techniques were used for isolation and purification of the principles. The structures were elucidated on the basis of spectral data (UV, IR, MS, (~1H-NMR), (~(13)C-NMR), (~1H-~1H COSY) and (~1H-~(13)C COSY), HMBC, and HMQC) and identified by comparing with the authentic substance. Results Five compounds were got from fat-soluble fraction. They are identified as: N-pentacosyl-2-carboxy-benzoyl amide (Ⅰ), 5-carboxyl-3, 4-dihydrogen-1H-2-benzopyran-1-one (Ⅱ), erythrocentaurin (Ⅲ), roburic acid (Ⅳ), oleanolic acid (Ⅴ). Conclusion Compounds Ⅰ and Ⅱ are two novel ones. They are named as qinjiaoamide and erythrocentauric acid, respectively. Compound Ⅲ is isolated from the plants of Gentiana L. for the first time.